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LA Pignataro, V Sarhty; Characterization Of Glutamate Transporters Expressed In Retinoblastoma Cells . Invest. Ophthalmol. Vis. Sci. 2002;43(13):728.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: L-Glutamate is the major excitatory neurotransmitter in the retina, and glutamate transporters are thought to play a role in regulating synaptic glutamate levels and neuronal damage following retinal ischemia. Four glutamate transporters, EAAT1, EAAT2, EAAT3 and EAAT5 have been found in the human retina. In order to facilitate gene regulation studies of the transporters using cell transfection assays, we have examined the profile of glutamate transporter expression in two well-established human retinoblastoma cell lines. Methods: Total RNA was isolated from cultures of the retinoblastoma cell lines, Y79 and WERI-Rb-1. Duplex relative reverse transcription-polymerase chain reaction (RT-PCR) analysis was performed using ribosomal (r)18S RNA as internal standard including competimer technologyTM. CCD camera images of ethidium bromide stained PCR products were quantified with the program Scion Image for Windows (SCION Corp, MD). EAAT optical density values were normalized to r18S RNA. Results: We found that EAAT1, 2, 3 and 5 were expressed in both Y79 and WERI-Rb-1 cells. However, there were quantitative differences. In Y79 cells, EAAT2 was expressed at a high level while EAAT1 and EAAT3 transcripts were less abundant. Surprisingly, EAAT4, not normally found in the retina, was detected in Y79 cells. In the WERI-Rb-1 cells, EAAT1, 2 and 3 were found at a high level. In contrast to Y79 cells, no EAAT4 mRNA was seen in WERI-Rb-1 cells. In the case of EAAT5, both cell lines showed a very high level of expression. EAAT5 mRNA level was more abundant than that of EAAT1, 2, 3 and 4 combined. Conclusion: These results indicate that although the two cell lines are derived from retinoblastoma tissue, they do not share the same profile of EAAT expression. Moreover, these cells which, have been used as models for cone photoreceptors, appear to express the glial glutamate transporter EAAT1. From the present study, it appears that Y79 and WERI-Rb-1 cells would be useful for molecular studies of EAAT5. Supported by NIH grants EY-03523, EY-07128 and EY-13125
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